Fluid separatory device

ABSTRACT

The device includes a barrel and plunger assembly for drawing the fluid into a separating chamber, chemical separating means such as an agglutinating agent for separating the phases in the chamber and means in the plunger assembly including a one-way valve for mechanical separation of the less dense supernatant phase from the more dense phase. The device is of particular utility in separation of plasma from whole blood in preparation for chemical analyses of the plasma.

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ABSTRACT: The device includes a barrel and plunger as- 20 NH 25 b 1 6 Al o H k F m 5 8 7 2 P 8 1i 2 M assembly including a one-way valve formechanical separation of the less dense supernatant phase from the moredense [56] References Cited UNITED STATES PATENTS 657,440 9/1900 McCawphase. The device is of particular utility in separation of plasma fromwhole blood in preparation for chemical analyses of the plasma.

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SHEET 1 BF 2 INVENTOR. James W Wmke/man flTTORNEKS FLUID SEPARATORYDEVICE BACKGROUND OF THE INVENTION 1. Field of the Invention Theinvention is directed to the separation of liquid phases from fluidmixtures. The invention is more particularly directed to separation ofparticular liquid phases from fluid mixtures for performing analyticalchemical operations on the liquid phases, and in a preferred embodiment,for separating plasma from blood. The term fluid mixture" hereinemployed to designate liquid compositions of a more or less homogeneouscharacter but which are separable into two or more separate phases ofdifferent densities so that such separation in a gravitational fieldresults ultimately in the disposition of the phases as layers, the moredense fluid phases being the lower layers and the less dense fluidphases forming the upper layers. The term fluid as employed with respectto mixtures and phases is intended to refer to solutions, liquidemulsions, colloidal dispersions, suspensions, slurries, suspensions ofcells such as blood cells, tissue homogenates or bacterial cultures andthe like.

2. Description of the Prior Art The separation of particular liquidphases from fluid mixtures has been carried out by a variety ofprocedures, one of the most widely used procedures involvingcentrifugation. In the separation of plasma from blood, for example, ablood sample is drawn from a subject, mixed with an anticoagulant andplaced in a centrifuge tube. The tube is placed in the centrifuge whichis counterbalanced for the weight of the sample and the sample iscentrifuged until the blood cells are sedimented, leaving the plasma asa supernatant fluid. The plasma can then be decanted or pipetted into asuitable container for storage, shipping or immediate clinical chemicalanalysis. The supernatant plasma is miscible with the more dense,cell-containing phase. The mechanical separation of the plasma requiresa high degree of caution on the part of a trained analyst to avoidturbulence in the mixture resulting in resuspension of cells in theplasma. Frequently it may be undesirable to expose the fluid to theatmosphere by transferring it from initial container, to centrifugetube, to final container as fluids such as blood can contain viruses ormicro-organisms which can create a biological hazard to the analyst.Likewise, the fluid to be separated can be contaminated during theseparation procedures so that certain analytical results can beaffected. Moreover, the separation requires additional equipment such asthe centrifuge, the additional tubes, etc. and decantation or pipettingof the supernatant fluid may require a skilled technician to preventremixing of the separated phases during the mechanical separationthereof. Moreover, in circumstances in which control of temperature isdesired during the separation, the operation is complicated by the needto either heat or cool the centrifuge, pipettes, tubes and otherauxiliary equipment.

Their is a need for a single simple device for separation of fluids inwhich separation of fluid mixtures into phases and mechanical separationof phases can be accomplished accurately in the same device whileminimizing turbulence and risks of remixing the phases.

SUMMARY OF THE INVENTION The present invention is concerned withapparatus useful for the separation of different fluid phases from afluid mixture. The invention is particularly directed to apparatususeful of the separation of fluid mixtures into phases of differingdensities and the mechanical separation of the phases from each other.

It is an object of the invention to provide a fluid-separatory device inwhich collection of fluid, phase separation and mechanical separation ofphases can be carried out in the same apparatus in a simple procedureinvolving a minimal number of distinct operations. It is a furtherobject of the invention to provide a fluid-separatory device which isinexpensive and simple in operation and construction, and which can beemployed as a container for shipping or storing the fluid phasesseparated therein. It is a further object of the invention to providea'device for the separation of a fluid phase such as plasma from wholeblood which is adapted to permit the entire operation to be carried outin a single vessel with minimum possibility of cross-contaminationbetween a technician using the device and the sample of blood and aminimum of crosscontamination between phases resulting from turbulenceat the interface. A further object is to provide a separatory devicewhich is adapted to operate under a variety of temperature conditionswithout requiring elaborate provisions for heating and cooling.

The apparatus includes a barrel having a plunger assembly movablydisposed therein, so that movement of the plunger unit varies the volumeof a sample chamber. The device includes inlet means for introducing afluid mixture into the chamber and chemical means in the chamber forfacilitating the separation of themixture into a plurality of phases ofdiffering densities. The device also includes means in the plunger unitfor mechanically separating the phases by permitting one phase to flowthrough the plunger unit as the plunger unit is moved to the interfaceof the two phases and preventing such separated phase from flowing backthrough the plunger unit toward the interface. The inlet means of thedevice can be coupled to a sampling element such as a hypodermic needleso that collecting the fluidmixture, separation into phases andmechanical separation of one phase from another are accomplished in thesame device. The device can also include spacer means which can beemployed in lie of or in combination with the chemical means tofacilitate the use of the apparatus in a centrifuge. The device issimple in construction and operation and capable of carrying outcomplete mechanical separation of fluid mixtures with a minimum ofcross-contamination between phases or between the fluid mixture and theatmosphere. It can be employed with or without a centrifuge and is thuseasily adapted to operation in a refrigerator or water bath or the like.

BRIEF DESCRIPTION OF THE DRAWINGS Other objects and advantages of heinvention will be apparent on consideration of the following descriptionand claims and of the drawings wherein:

FIG. 1 is an isometric view of a fluid-separatory device of theinvention with a portion 'ofthe barrel and plunger piston broken away toshow the chemical separatory means;

FIG. 2 is a cross-sectional view of the device of FIG. 1 taken alongline 2-2 of FIG. 1 and showing the device in one state of use;

FIG. 3 is a cross-sectional view of the device of FIG. 1 taken alongline 2-2 and showing the device in another state of use;

FIG. 4 is a cross-sectional view of the device of FIG. 1 taken alongline 2-2 and showing the device in a third state of use.

DESCRIPTION OF THE PREFERRED EMBODIMENT Referring more particularly tothe drawings, the separating apparatus 10 comprises a barrel 12 havingside and end walls l4, 16; a plunger member 20, including a plungerpiston 22 slidably disposed in the barrel in fluidtight relation to thesidewall 14 thereof so that the plunger piston 22 and the walls 14, 16of the barrel define a fluidtight separating chamber 40 of variablevolume; disengageable spacer means 70 for selectively limiting movementof the plunger piston 22 in the barrel at a predetermined-point wherebya predetermined minimum volume of chamber) is defined by the piston 22and walls 14, 16; closeable inlet mearts 18 on the barrel 12 forintroducing fluid into the chamber 40; chemical means 42 forfacilitating the separation ofa fluilmixture into two fluid phases, saidmeans being disposedii'n the chamber 40; means for so disposing thechemical means 42 in chamber 40; and conduit means 24 in the plungerpiston 22 for providing unidirectional fluid flow from chamber40 throughthe plungerpis ton 22.

The term fluidtight as herein employed in reference to the relationshipof the plunger piston 22 and the barrel sidewall 14 means and refers tothat degree of fit of the plunger piston 22 within the barrel whichpermits fluid to enter or leave the camber 40 only through inlet means18 or conduit means 24. Such fit permits fluid to be drawn into chamber40 through the inlet means 18 against the force of gravity by thesuction created by movement of the plunger piston away from the end wall16 of the barrel 12. The fluidtight relation of the plunger piston andthe barrel thus corresponds to the relation of the piston and cylinderof a conventional lift pump or the relation of the plunger and syringebarrel of a conventional hypodermic syringe.

The barrel 12 can have any configuration or size compatible with theslidable disposition of the plunger piston 22 therein. A cylindricalconfiguration similar to that of a syringe barrel is preferred. Thedimensions of the barrel are conveniently such as to provide a samplechamber 40 with a maximum volume of from about 5 to about 50 to 100cubic centimeters. A barrel corresponding to the syringe barrel of aconventional or cubic centimeter hypodermic syringe is convenient. Thebarrel can likewise be of any suitable material which is sufficientlyrigid and fluid resistant to contain the fluids to be separated. Glass,metal, hard rubber or synthetic plastic materials are representativematerials. It is greatly preferred that the barrel 12 be transparent, sothat movement of the plunger piston 22 therein and separation of fluidphases can be easily observed. Consequently, it is greatly preferredthat the barrel be constructed of a transparent material such as glass,or a transparent plastic such as polyethylene, polypropylene orpolystyrene.

The closeable inlet means 18 is preferably disposed in the end wall 16of the barrel 12 and comprises an opening or port through the end wall16 to provide communication between the inside and outside of thebarrel. In a preferred embodiment, the inlet means 18 also includes atubular nipple 19. The nipple 19 is adapted to be coupled to afluid-sampling device 36 by friction fit of an adapter 38 about thenipple. Sampling device 36 can be a conventional hollow hypodermicneedle. Alternately, the adapter 38 and sampling device or needle 36 canbe removed and a fluidtight cap 32 can be releaseably secured over thenipple 19 by friction fit as shown in FIG. 3, thus selectively closingthe inlet means. The inlet means must be capable of closure to preventflow of fluid from sample chamber 40 at certain times during theoperation of the device, although the means by which closure isaccomplished are not critical. Thus, other means may be employed toprovide for selectively closing the inlet means 18. For example, aresilient plug can be inserted into the inlet means, or the nipple canbe coupled to a sampling device by means ofa resilient tubular adapterwhich can be closed by a clamp or the like. Also, the inlet means 18 canhave a one-way valve permitting fluid to enter chamber 40 through theinlet means 18, but preventing the exit of fluid through the inletmeans.

The plunger unit 20 and piston 22 can have any configuration whichcorresponds to the slidable fluidtight disposition of the plunger piston22 in the barrel 12. The plunger unit 20 can also include a plunger rod21 which extends upwardly from the piston 22 and away from the end wall16 of the barrel. Preferably, the plunger piston 22 and plunger rod 21are formed as a unit from the same type of material employed in themanufacture of the barrel. The combined length of both plunger elementsshould be such that a portion of the plunger rod 21 extends beyond thesidewall 14 of the barrel 12 when the plunger piston 22 is disposedagainst the end wall 16 of the barrel. Both the barrel l2 and theplunger rod 21 can have manually graspable protrusions 60 extendingoutwardly therefrom to facilitate manual control of the sliding motionof the plunger piston 22 in the barrel 12.

The apparatus 10 can also include a disengageable spacer 70 adapted tofit between a protrusion 60 on the barrel and a corresponding protrusion60 on the plunger rod 21 as shown in FIG. 2. The spacer 70 can be of anysuitable material of sufficient rigidity to maintain the plunger rod andpiston 21, 22 in a predetermined spaced relation from the end wall 16 ofbarre] 12 when the apparatus is employed in a centrifuge. When 70 isengaged with protrusions 60 as shown in FIG. 2, the downward motion ofplunger piston 22 is limited so that the volume of chamber 40 cannotdecrease below a predetermined limit. The dimensions of space 70 areselected so that the predetermined minimum volume of chamber 40 is atleast as great as the volume of fluid mixture desired to be separated.When the apparatus and fluid mixture are centrifuged, the spacer 70prevents the plunger piston 22 from moving under centrifugal force. Asecond disengageable spacer adapted to provide a second predeterminedvolume to chamber 40 can also be employed. Such second volume is equalto or preferably slightly greater than the volume of the more densefluid to be retained in chamber 40 after the less dense supernatantfluid is mechanically removed. A single rectangular plate having alength and breadth corresponding to the two predetermined volumes,respectively, is conveniently employed as a spacer.

The unidirectional conduit means 24 extends through the piston 22 andincludes a one-way check vale 2. The exact type of one-way check valveto be employed is not critical and a ball valve, butterfly valve,spring-biased valve or the like can be employed. However, it isessential that the valve 26 and conduit be such as to permit fluid toflow upward through the plunger piston 22 from chamber 40 and to preventany flow of fluid downward through the piston 22 into chamber 40. Aone-way, gravity-biased ball valve gives excellent performance inproviding the required unidirectional flow, and a ball valve is thepreferred one-way valve. In the valve 26, a ball 25 is disposed in avalve chamber 27 which comprises an enlarged portion of the conduit 24.In the upper portions of the valve chamber 27, a stop 29 extends intochamber 27. Stop 29 is so disposed in the upper portion of chamber 27 asto prevent the ball 25 from blocking fluid flow from chamber 27 upward,and is of such dimensions as to permit upward flow even when the ball isin its uppermost position. When the device 10 is in its normal verticalposition in which the barrel end wall 16 is below the plunger piston 22,the ball 25 is biased by gravity into a position blocking the conduitmeans as shown in FIGS. 1 and- 2. Suction created in chamber 40 as itsvolume is increased by upward sliding of the plunger piston 22 andincreasing the volume of chamber 40 as shown in FIG. 2, or the weight ofany fluid disposed in the barrel 12 above the piston 22 as shown in FIG.4 provides an additional bias toward closure of the ball valve. Theupward pressure of any fluid in chamber 40 results in upward and/orlateral displacement of the ball 25, opening valve 26 to permit fluidflow through the conduit means 24 as illustrated in FIG. 3. Downwardsliding motion of the plunger piston 22 in the barrel 12 when the inputmeans 18 are closed increases fluid pressure in chamber 40 as the volumeof chamber 40 decreases and results in opening the valve 26. Thus, theone-way valve 26 is adapted to permit unidirectional flow from thechamber 40 through the valve when the plunger piston is moved downwardor toward end wall 16.

The device 10 preferably includes receptacle means for collecting fluidwhich passes from chamber 40 through the plunger piston 22. In apreferred embodiment shown in the drawings, the plunger rod 21 extendsupwardly from the plunger piston 22 and entirely surrounds the conduitmeans 24, whereby the plunger piston 22 and rod 21 define a fluidreceptacle or chamber 80. Receptacle is thus disposed downstream of heplunger piston 22 and conduit means 24 with respect to unidirectionalflow of fluid therethrough. Receptacle or chamber 80 is closed atanother end by an end wall 33 which can be a portion of the plunger rod21. End wall 33 includes a sealable aperture 30 surrounded by a nipple31 extending outwardly from end-wall 33. A removable sealing cap 34 canbe releasably secured over nipple 31 to seal the aperture 30 as shown inFIGS. 1 and 2. With the cap 34 over the nipple, the pressure of air inreceptacle 80 resists the flow of fluid through conduit 24 and permitsthe device 10 to be handled easily without entry of fluid intoreceptacle 80 until such flow is desired. The cap 34 can be removed whendesired to permit air enclosed in fluid receptacle 80 to escape to theatmosphere through aperture 30 as fluid passes through conduit means 24into receptacle 80. Tl-lus, the aperture 30 and cap 34 provideadjustable means for relief of pressure in the receptacle 80 bypermitting selective venting to the atmosphere. The plunger piston 22,plunge rod 21 and receptacle 80 included therein, and the end wall 33are preferably formed as a single plunger unit which can be separatedfrom the barrel when a fluid phase sample has been collected inreceptacle 80. On separation of the plunger unit 20 from barrel 12 afterusing the device, the device serves as a pair of separate containers forseparated fluids. Conventional means can be used for sealing conduit 24after separation of the plunger unit and barrel such as a plug (notshown) adapted to fit into the conduit by friction fit, screw threads orthe like, or a cap (not shown) adapted to fit over the lower end of theplunger assembly in substantially the same manner as cap 34 fits overnipple 31.

The chemical means 42 disposed in chamber 40 can be disposed therein inany fashion which will ensure that the chemical means act on the fluidmixture to be separated and to separate the same into two phases. Thus,the chemical means can be present as a solid or a liquid in chamber 40,for example, as a liquid film or a solid coating on the interior of thebarrel sidewall 14 or on the surface of the plunger piston 22 or thebarrel end wall 16, in which case the film or coating serves as meansfor introducing the chemical separating means into a fluid mixturedisposed in chamber 40. The film or coating is applied to the piston orinterior of the barrel by removing the plunger unit from the barrel,applying the chemical means and replacing the plunger unit in thebarrel. Alternately, it can be present as a mixture with thefluid to beseparated, having been mixed with such fluid either prior to orsubsequent to the placing of the fluid in the device 10, in which casethe inlet 18 serves as means for introducing the chemical separatingmeans into chamber 40 and thus into contact with the fluid mixture. Itis greatly preferred, however, that the chemical means be disposed inchamber 40, whether or not a fluid to be separated is simultaneouslypresent.

The chemical means can be any agent suitable for enhancing separation ofthe particular liquid mixture into two separate phases. When the liquidis an aqueous emulsion such as emulsions of various organic liquids andwater, or of fats or oils and water, the chemical means can be anemulsion-breaking agent. When it is desired to separate colloidaldispositions or protein suspensions or solutions into supernatant liquidand a liquid phase containing solid material, a flocculant or a proteinprecipitant can be employed. When the liquid mixture is a suspension ofcells such as blood, aqueous bacterial cultures or tissue homogenates,the chemical agent can be agglutinant, that is, an agent which enhancesthe agglutination or clumping together of cellular material. Inparticular applications, selective chemical separating agents capable ofseparating particular liquid mixtures into particular fluid phases canbe employed; for example, when the liquid mixture is milk, anemulsion-breaking agent such as amyl alcohol can be employed to separatethe fat from the heavier, aqueous colloidal phase or a proteinprecipitant such as silver nitrate or lead acetate can be employed toseparate the milk into a proteinfree liquid phase and a liquid phasecontaining precipitated flocculated protein. The chemical separatingmeans cooperates with the other elements of the device to facilitate thecomplete separation of the desired fluid phase and, in many cases, thechemical separating means determines-the nature of the supernatant fluidseparated from the fluid mixture. The exact chemical separating means tobe employed is dependent upon the particular fluid mixture to beseparated by the use of the device and the particular liquid phases tobe obtained, it being essential only that the chemical means facilitatethe separation of the liquid mixture into two or more separate liquidphases of different specific gravities.

Various chemical separating agents which can be employed in theseparation of particular fluid mixtures are disclosed by Clayton, Theoryof Emulsions,5th Edition, chapter Xlll, Blakiston Company, New York(I954) and by Becher, Emulsions: Theory and Practice, Second Edition,chapters 5 and 9, ReinholdQNew York (i965). Agents useful inprecipitation of colloidal materials or suspensions are disclosed byShaw, introduction to Colloid and Surface Chemistry, chapters 8 and 10,Butterworths, London (1966). Chemical means useful in the separation ofbiological fluid phases in fluid mixtures such as blood are disclosed byHenry, Clinical Chemistry: Principles and Techniques, Hoeber Division ofHarper and Rowe (Second Printing, 1964). I

The device of theinvention has particular utility in the separation ofplasma or serum from whole blood, typically in preparation for chemicalanalysis of various plasma or serum components. In such use, thechemical separating means 42 comprises an agglutinant and, optionally,an anticoagulant, and preferably both an agglutinant and ananticoagulant. The agglutinant serves to bring about the clampingtogether and precipitation of the blood cells and to separate the bloodinto a cell-free liquid phase and a more dense liquid phase containingthe precipitated clumps of cells. Representative agglutinants which canbe employed as chemical separating means include fibrinogen,polyvinylpyrrolidone, hemagglutinin, phytohemagglutinin and dextran. Thepreferred agglutinant is fibrinogen. When an agglutinant is the solechemical-separating means, clotting of blood takes place as well asagglutination. In such a case,'serum is obtained as the less denseliquid phase above the precipitated cells. Separation of serum ratherthan plasma is desirable in certain situations. In other cases, plasmais desired, and the loss of plasma components such as prothrombin whichare involved in clot formation is undesirable. Thus, in order toseparate plasma from blood in the device of the invention, the chemicalagent must comprise an anticoagulant, that is, a chemical agent whichinhibits the clotting of blood, as well as the agglutinant required toseparate the blood cells into a separate liquid phase. Representativeanticoagulants which can be employed include heparin and syntheticheparinlike compounds; bishydroxycoumarin; oxalate anticoagulants suchas potassium oxalate or ammonium oxalate; citrate anticoagulants such astrisodium citrate, tripotassium citrate and mixtures of trisodiumcitrate, citric acid and dextrose; alkali metal salts ofethylenediaminetetraacetic acid; potassium fluoride and the like.Heparin is the preferred anticoagulant.

Either the anticoagulant or the agglutinant or both can be disposed inthe chamber 40 prior to the introduction'of blood thereinto. Preferably,both the anticoagulant and the agglutinant are disposed as a solid layeror deposit on an internal wall of barrel 12 or the plunger piston 22,and disposition of a solid mixture of the two agents on the surface ofend wall 16 is preferred. The chemical means 42 for separation of plasmafrom blood is conveniently disposed in chamber 40 by withdrawing theplunger unit 20 from the barrel, pouring a solution of heparin andflbrinogen into the barrel and allowing the solvent to evaporate,leaving the mixture of anticoagulant and agglutinant as a solid layer onend wall 16.

Tile amount-of chemical separating means to be employed depends in parton the amount of fluid mixture to be separated and on the result to beachieved. THe chemical separating agent should be employed in an amountsuch that the dilution and admixture of the agent with the particularfluid mixture provides an effective concentration of the agent in themixture. An effective concentration is sufficient to produce the desiredeffect, emulsion breaking, precipitation, clot inhibition, agglutinationor the like. In plasma separation for example, the chemical means 42includesesufiicient heparin to provide an anticoagulant concentration offrom about 10 to about 20 to about 50 milligrams of heparin permilliliters of blood, and sufficient fibrinogen to provide anagglutinating amount of from about 50 to'about 200 to about 600milligrams of fibrinogen per 100 milliliters of blood.

In the operation of the device 10, a fluid mixture 44 to be separated isdrawn into chamber 40 through the inlet means 18 as the plunger piston22 is slidably moved away from barrel end wall 16. Inlet means 18 isthen closed by cap 32 and the fluid mixture 44 is mixed with thechemical-separating means 42 which has been disposed in chamber 40 priorto introduction of fluid mixture 44. The device 10 is then maintained ina substantially vertical position as shown in FIG. 2 until the fluidmixture 44 separates into a less dense supernatant fluid phase 52,situated in the upper portion, and a more dense fluid phase 54, situatedin the lower portion of chamber 40. Alternately, a spacer 70 is engagedbetween protrusions 60 on the barrel 12 and plunger rod 21 to maintainthe minimum volume of chamber 40 at least equal to the volume of fluidmixture 44, and the device 10 is centrifuged by conventional proceduresto dispose the more dense phase 54 in that portion of-chamber 40 nearestend wall 16. Depending upon the fluid mixture employed and the type ofseparation desired, the liquid phases can be separated by centrifugationwhile the spacer is engaged without employing a chemical-separatingmeans. The cap 34 is then removed from aperture 30 and when a spacer 70has been employed, the spacer is disengaged or removed. The plungerpiston 22 is then slidably moved downward toward barrel end wall 16. Asplunger piston 22 moves into the chamber 40, the upper less dense fluidphase 52 passes through unidirectional conduit 24 and valve 26 into thefluid receptacle 80 as shown in FIG. 3. Downward motion of the plungerpiston 22 is continued until the plunger piston 22 reaches the interface50 between the two liquid phases. Alternately, a second spacer isengaged with a protrusion 60, the second spacer being adapted to preventdownward motion of the plunger piston beyond the interface 50, and theplunger piston 22 is moved downward until further downward motion isprevented by the spacer. Since the less dense liquid in receptacle 80 isprevented from returning to chamber 40 by oneway valve 26, mechanicalseparation of the two liquid phases 52, 54 is complete at this stage anthe plunger unit can be moved upward as shown in FIG. 4. If desired, cap32 can be removed from the barrel nipple 19 and the more dense fluid 54can be drained from chamber 40 through the inlet means 18. Also, theplunger unit 20, including the less dense liquid phase 52 disposed inreceptacle 80, can be removed and aperture 30 can be capped and conduit24 closed in any convenient manner to provide a closed receptacle forstoring or handling the fluid phase 52. Since the device can be employedwithout centrifugation, the temperature at which separation is carriedout can be controlled easily by placing the device in a refrigerator,incubator, ice bath, water bath or the like.

In the separation of plasma from whole blood, the chemical means 42employed is preferably a solid film of heparin and fibrinogen, and themixture of the anticoagulant and the agglutinant is disposed as a filmon end wall 16 as shown in FIG. 1. The chemical means is deposited onthe end wall as described above, prior to using the device. The deviceequipped with a needle 36 and adapter 38 on inlet means 18 and a cap 34on aperture 30 is employed to draw a sample of blood from a subject intochamber 40. The device 10 is inverted, the needle and adapter 36, 38 areremoved and cap 34 is placed on inlet means 18. With the inlet meansthus closed, the anticoagulant and agglutinant are mixed with the bloodfluid mixture 44 by inverting the apparatus several times. The device isthen placed in an upright position as shown in FIGS. 2-4 and held for ashort period while the plasma separates from the blood cells as an upperfluid phase 52. Generally, the separation is sufficiently rapid so thatplasma occupies the upper 45 to 55 percent of the sample chamber volumeafter about 30 minutes or the upper 50 to 60 percent about 1 hour. Whenit is desired to separate serum or plasma from whole blood by employingthe device in conjunction with a centrifuge, a spacer 70 is employed asdescribed above. When a centrifuge and spacer are employed in separatingplasma, the chemical separating means can comprise an anticoagulantalone. With serum, rather than plasma, is separated in this procedure,the chemical means can be omitted or can comprise an agglutinant alone.After the phases have separated to the desired extent, cap 34 is thenremoved from aperture 30 and the spacer 70, if used, is disengaged. Theplunger piston 22 is then urged downward toward the interface 50 of theplasma phase and the blood cell phase. The plasma flows through theconduit means 24 into receptacle as shown in FIG. 3. If desired, theplunger piston can be moved slowly downward during the holding period toprovide for gradual continuous flow of plasma through conduit 24 toreceptacle 80 as the separation of phases by agglutination of the cellsprogresses. When the plunger piston 22 reaches the interface 50 or apoint slightly above it, the plunger unit 20 is withdrawn upwardly fromthe barrel 12 as'shown in FIG. 4. 'Cap 34 is replaced on nipple 31 and,if desired, conduit means 24 can be sealed by a plug or the like. Theplasma separated in the device can be shipped or stored in thereceptacle 80 and is suitable for analytical clinical chemical purposes.

It will thus be seen that the device can be described as an improvedsyringe similar to conventional hypodermic syringes. Thus, the device 10has a barrel l2 equipped with a detachable needle 36 and a plunger unit20 including a plunger piston 22 and plunger rod 21. In the device ofthe invention, however, the plunger piston includes conduit means 24 forproviding unidirectional flow from the barrel into a receptacle 80enclosed within the rod 21 and means for selectively preventing loss offluid from the barrel 12 such as the cap 34 over inlet means 18.Moreover, the device 10 includes chemical separation means 42 in chamber40 of the barrel 12 and means such as inlet 18 or the removable plungerunit for disposing chemical means 42 in chamber 40. Thus, the devicecorresponds to a syringe which is adapted to bring about the separationof fluid mixtures disposed .therein by chemically enhanced separation ofthe fluid mixture into phases and also by mechanical separation of oneliquid phase from another.

The device can be modified, for example, by the choice of particularunidirectional flow means in the plunger piston, the choice ofparticular closeable inlet means on the barrel or means of closing theaperture on the receptacle in the plunger unit. Modification of the'chemical-separating means by the choice of particular means toaccomplish particular functions in the separation of particular fluidmixtures can be carried out to adapt the device for use in a variety ofseparatory applications. Also, it will be apparent that in some.applications, one or more spacers can be employed to permitcentrifugation of the device and omission or variation of the chemicalseparating means. Mechanical means for moving the plunger piston canalso be employed, with such means being controlled by timers or sensingmeans such as photoelectric devices for actuating the mechanical meanswhen a predetermined amount of supernatant fluid has separated from thefluid mixture.

What I claim is:

1. A method for separating a fluid mixture into a plurality of fluidphases of differing densities within a syringe having a barrel,including a sidewall and an end wall, inlet means in a wall of thebarrel for admitting a fluid mixture thereto; a plunger slidablydisposed in the barrel in fluidtight relation to the sidewall thereof soas to form a chamber of variable volume in the barrel, means forselectively closing the inlet means; conduit means on the plunger forproviding unidirectional fluid flow through the plunger from thechamber; and a receptacle on the plunger surrounding the conduit meansand downstream thereof, the method comprising:

a. drawing a fluid mixture to be separated through the inlet means andinto the chamber,

b. mixing the fluid mixture in the chamber with a chemical means forfacilitating the separation of a fluid mixture into a plurality of fluidphases of differing densities,

c. closing the inlet means, and thereafter d. holding the syringe in avertical position with the plunger above the defined chamber until thefluid mixture separates into a plurality of phases of differingdensities,

e. moving the plunger downward into the chamber toward the interfaceresulting from separation of the mixture and simultaneously venting thereceptacle to atmosphere, whereby the uppermost liquid phase flowsthrough the conduit means into the receptacle;

f. terminating the downward motion at a point in said chamber not belowthe interface;

g. and thereafter withdrawing the plunger, receptacle and the liquidphase therein from the barrel.

2. The method of claim 1 further comprising the step of sealing thereceptacle from co mmunication with atmosphere 22;;3?" UNITED STATESPATENT OF'FICE CERTIFICATE OF CORRECTION Patent No. 3.596,652 Dated 3August 1221 Inventor(s) James W. Winkelman It is certified that errorappears in the above-identified patent and that said Letters Patent arehereby corrected as shown below:

In column 2, line 32, change "lie" to lieu Column 5, line 26, delete"liquid in" and insert liquid disxxjsedg. in"-; in line U8, delete"dispositions" and insert dispersions Column 6, line 21, delete"clamping" and insert clumping Signed and sealed this 11th day of April1972.

(SEAL) Attest:

EDWARD M.FLETCHER JR. Attesting Officer ROBERT GOT'ISCHALK Commissionerof Patents

1. A method for separating a fluid mixture into a plurality of fluidphases of differing densities within a syringe having a barrel,including a sidewall and an end wall, inlet means in a wall of thebarrel for admitting a fluid mixture thereto; a plunger slidablydisposed in the barrel in fluidtight relation to the sidewall thereof soas to form a chamber of variable volume in the barrel, means forselectively closing the inlet means; conduit means on the plunger forproviding unidirectional fluid flow through the plunger from thechamber; and a receptacle on the plunger surrounding the conduit meansand downstream thereof, the method comprising: a. drawing a fluidmixture to be separated through the inlet means and into the chamber, b.mixing the fluid mixture in the chamber with a chemical means forfacilitating the separation of a fluid mixture into a plurality of fluidphases of differing densities, c. closing the inlet means, andthereafter d. holding the syringe in a vertical position with theplunger above the defined chamber until the fluid mixture separates intoa plurality of phases of differing densities, e. moving the plungerdownward into the chamber toward the interface resulting from separationof the mixture and simultaneously venting the receptacle to atmosphere,whereby the uppermost liquid phase flows through the conduit means intothe receptacle; f. terminating the downward motion at a point in saidchamber not below the interface; g. and thereafter withdrawing theplunger, receptacle and the liquid phase therein from the barrel.
 2. Themethod of claim 1 further comprising the step of sealing the receptaclefrom communication with atmosphere during the mixing and holding steps.3. THe method of claim 1 wherein the chemIcal means for facilitatingseparation comprises an agglutinant.
 4. THe method of claim 3 whereinthe chemical means for facilitating separation further comprises ananticoagulant.
 5. THe method of claim 4 wherein the anticoagulant isheparin and the agglutinant is fibrinogen.